The Vpu protein is a small membrane bound protein that has been shown to: a) down-modulate CD4 from the cell surface by shunting CD4 to the proteasome pathway; b) facilitate the release of virions from the infected cell; and c) have an ion channel activity. A major drawback in studying the role of Vpu in lentiviral pathogenesis has been lack of a vpu gene in simian immunodeficiency virus (SIV), which has been the most commonly used macaque model to study AIDS pathogenesis. As an alternative to the SIV/macaque model, chimeric simian-human immunodeficiency viruses (SHIVs) have been developed that contain the tat, rev, vpu, and env genes from HIV-1 in a genetic background of SIVmac239 and have been used to derive pathogenic variants that when inoculated into pig-tailed or rhesus macaques, cause high virus burdens, massive CD4+ T cell loss and AIDS, and death within 6 months to 1 year. Studies from our laboratory have indicated that the Vpu does play a role in the CD4+ T cell loss caused by these viruses. We have recently developed a Vpu/enhanced green fluorescent protein fusion protein reporter system (VpuEGFP) that mimics the intracellular transport, intracellular stability and CD4 down-regulation of Vpu protein within virus infected cells. In this proposed studies, we will use a recently developed VpuEGFP reporter system to: a) assess the role of the different Vpu domains on the retention of subtype B Vpu in the Golgi complex; b) to use site-directed mutagenesis to analyze the role of the structure-function relationships of the amino acids comprising the cytoplasmic domain of the subtype B Vpu; and c) to compare the biological properties with the divergent Vpu proteins from subtype C isolates of HIV-1. Finally, we will introduce Vpu proteins having altered biological properties in a molecular clone of SHIV to assess the role of the different biological properties of Vpu on CD4+ T cell loss caused by SHIV in pig-tailed macaques. The proposed studies will provide new information on the structure-function relationships of the different domain s of Vpu and the role of the different biological properties of Vpu on HIV-1 pathogenesis.